Interaction of sphingomyelins and phosphatidylcholines with fluorescent dehydroergosterol.
Interaction of sphingomyelins and phosphatidylcholines with fluorescent dehydroergosterol.
复制标题
鞘磷脂和磷脂酰胆碱与荧光脱氢麦角甾醇的相互作用。
DOI：
10.1021/bi00440a041
复制
发表时间：
1989
期刊：
影响因子：
2.9
通讯作者：
Nemecz,G
中科院分区：
文献类型：
--
作者：
Schroeder,F;Nemecz,G
药理学和药物化学系，药学院，药理学和细胞生物物理学系，医学院，辛辛那提大学医学中心，3223 Eden Avenue，辛辛那提，俄亥俄州45267-0004接收日期：1988年8月24日;修订后的手册接收日期：1989年4月3日摘要：将荧光甾醇脱氢麦角甾醇作为胆固醇类似物与多频相位和调制相结合（1-250 MHz）荧光测定法，以检查甾醇（1）是否优先与流体或固相磷脂相互作用，以及（2）是否优先与相分离或相分离的鞘磷脂相互作用。混溶性共声处理磷脂膜。通过将脂质在有机溶剂中混合、干燥混合物、加入缓冲液、超声处理和分离SUV来产生超声处理的小单层囊泡（SUV）。利用合成的以及生物来源的磷脂。在二肉豆蔻酰磷脂酰胆碱/二硬脂酰磷脂酰胆碱（DMPC/DSPC，摩尔比1：1）的相分离共超声SUV中，荧光甾醇（0.5mol%）优先与液相脂质相互作用（分配系数K（u=2.6-3.4））。首先，脱氢麦角甾醇仅检测到DMPC的相变，DMPC是具有较低相变温度的磷脂。第二，脱氢麦角甾醇的荧光偏振，极限各向异性，序参数，和旋转弛豫时间在共声处理囊泡是类似的脱氢麦角甾醇在SUV中仅由DMPC组成。第三，共超声SUV中脱氢麦角固醇荧光寿命组分的数量及其分布与DMPC组成的SUV中脱氢麦角固醇的数量及其分布相似。第四，在比DMPC SUV低得多的脱氢麦角甾醇浓度下，在DSPC SUV中检测到脱氢麦角甾醇浓度依赖性自猝灭。通过监测单独超声处理的DMPC SUV和DSPC SUV之间的脱氢麦角甾醇交换，也观察到脱氢麦角甾醇对液相脂质的偏好，在两种类型的囊泡以相等比例混合后。在这些SUV混合物中，脱氢麦角甾醇也分配到更流动的SUV中，99：1。与此相反，在largelyphase混溶，cosonicated DMPC/DPPC SUV，脱氢麦角甾醇的荧光特性并没有表明一个优先与磷脂的相互作用。在各种相分离的，共超声处理的鞘髓鞘/磷脂酰胆碱SUV或合成的（棕榈酰油酰磷脂酰胆碱/硬脂酰鞘磷脂和棕榈酰油酰磷脂酰胆碱/棕榈酰鞘磷脂）或生物（卵磷脂-磷脂酰胆碱/牛脑鞘磷脂）来源，荧光甾醇也优先与具有较低相变温度的脂质、棕榈酰油酰磷脂酰胆碱（K（s）大于13）或卵磷脂（K（s）= 13）相互作用。
Department of Pharmacology and Medicinal Chemistry, College of Pharmacy, and Department of Pharmacology and Cell Biophysics, College of Medicine, University of Cincinnati Medical Center, 3223 Eden Avenue, Cincinnati, Ohio 45267-0004 Received August 24, 1988; Revised Manuscript Received April 3, 1989 abstract: The fluorescent sterol dehydroergosterol was used as a cholesterol analogue in conjunctionwith multifrequency phase and modulation (1-250 MHz) fluorometry to examine whether sterols (1) interact preferentially with fluid-or solid-phase phospholipids and (2) interact preferentially with sphingomyelin in phase-separated or phase-miscible cosonicated phospholipid membranes. Cosonicated small unilamellar vesicles (SUV) were produced by mixing lipids in organic solvents, drying the mixture, adding buffer, sonicating, and separating SUV. Phospholipids of synthetic as well as biological origin were utilized. In phase-separated, cosonicated SUV of dimyristoylphosphatidylcholine/distearoylphosphatidylcholine (DMPC/DSPC, 1: 1 molar ratio), the fluorescent sterol (0.5 mol%) interactedpreferentially with the fluid-phase lipid (partition coefficient, K (u=2.6-3.4) accordingto four criteria. First, dehydroergosterol detected only the phase transition of DMPC, the phospholipid with the lower phase transition temperature. Second, the dehydroergosterol fluorescencepolarization, limiting anisotropy, order parameter, and rotational relaxation time in the cosonicated vesicle were similar to those of dehydroergosterol in SUV composed only of DMPC. Third, the number of dehydroergosterol fluorescence lifetime components as well as the distribution in the cosonicated SUV was similar to that of dehydroergosterol in SUV composed of DMPC. Fourth, dehydroergosterol concentration-dependent self-quenching was detected in DSPC SUV at much lower dehydroergosterol concentration than in DMPC SUV, Preference of dehydroergosterol for fluid-phase lipids was also observed by monitoring dehydroergosterol exchange between individually sonicated DMPC SUV and DSPC SUV after the two types of vesicles were mixed in equal proportions. In these SUV mixtures, the dehydroergosterol also partitioned into the more fluid SUV, 99: 1. In contrast, in largelyphase-miscible, cosonicated DMPC/DPPC SUV, the fluorescence properties of dehydroergosterol did not indicate a preferential interaction with either phospholipid. In a variety of phase-separated, cosonicated sphingo-myelin/phosphatidylcholine SUV of either synthetic (palmitoyloleoylphosphatidylcholine/stearoylsphingomyelin and palmitoyloleoylphosphatidylcholine/palmitoylsphingomyelin) or biological (egg phos-phatidylcholine/bovine brain sphingomyelin) origin, the fluorescentsterol also interacted preferentiallywith the lipid with lower phase transition temperature, palmitoyloleoylphosphatidylcholine (K {/s greater than 13) or egg phosphatidylcholine (Kfjs=